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WO2015170790A1 - Composition de traitement et de prévention des lésions ischémiques - Google Patents

Composition de traitement et de prévention des lésions ischémiques Download PDF

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Publication number
WO2015170790A1
WO2015170790A1 PCT/KR2014/004194 KR2014004194W WO2015170790A1 WO 2015170790 A1 WO2015170790 A1 WO 2015170790A1 KR 2014004194 W KR2014004194 W KR 2014004194W WO 2015170790 A1 WO2015170790 A1 WO 2015170790A1
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WIPO (PCT)
Prior art keywords
reperfusion injury
injury
ischemic
ischemia reperfusion
composition
Prior art date
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PCT/KR2014/004194
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English (en)
Korean (ko)
Inventor
김상재
양재석
김관민
전상훈
장지은
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Kael-GemVax Co Ltd
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Kael-GemVax Co Ltd
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Priority to PCT/KR2014/004194 priority Critical patent/WO2015170790A1/fr
Priority to CN201480080217.6A priority patent/CN106659149B/zh
Priority to KR1020167031602A priority patent/KR102232320B1/ko
Priority to ES14890834T priority patent/ES2962532T3/es
Priority to EP14890834.6A priority patent/EP3138399B1/fr
Priority to JP2016565207A priority patent/JP6466971B2/ja
Priority to PCT/KR2014/004752 priority patent/WO2015167067A1/fr
Priority to US15/307,632 priority patent/US10662223B2/en
Publication of WO2015170790A1 publication Critical patent/WO2015170790A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/10—Peptides having 12 to 20 amino acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof

Definitions

  • the present invention relates to a composition for treating and preventing ischemic injury. More specifically, the present invention relates to a composition comprising a peptide derived from telomerase, which is effective for treating and preventing ischemic injury.
  • Ischemic damage occurs when blood circulation is blocked in organs that require blood flow, including the heart, brain, kidneys, and lungs (acute infarction, cerebral infarction, and nerve infarction). Height can be called tissue damage. Ischemic injury causes fatal complications in the heart, brain, kidneys and lungs, and delays the transplantation recovery, increases acute rejection and reduces long-term survival.
  • ischemia a condition known as hypoxia.
  • hypoxia a condition known as hypoxia.
  • obstructive vascular disease CAD
  • coronary thrombosis CAD
  • cerebrovascular thrombosis CAD
  • aneurysm rupture CAD
  • systemic bleeding CAD
  • crush injury CAD
  • sepsis severe skin burns
  • Vascular occlusion surgery methods eg, spinal cord ischemia in thoracoabdominal aneurysm surgery
  • cardiopulmonary bypass methods organ transplantation
  • cardiopulmonary collapse acute heart death
  • reperfusion injury Additional damage caused by restoring blood flow and oxygen delivery is known as reperfusion injury.
  • Paradoxical tissue damage caused by reperfusion injury appears to be similar to acute inflammatory conditions resulting from the attachment of inflammatory cells to reperfused tissues, the activation of these inflammatory cells and the subsequent formation of free radicals [Granger et al. Ann. Rev. Physiol., 57, 311-332, (1995)].
  • the production of free radicals and other cytotoxic biomolecules in reperfused tissue can lead to cell death by necrosis or activation of apoptosis pathways.
  • Ischemic tissue damage caused by ischemia-reperfusion (IR) during organ transplantation results in delayed recovery of organ function after organ transplantation, which is a poor prognosis for long-term functioning of organs transplanted with inflammatory tissue reactions. Often act as a factor. Early ischemic reperfusion injury that occurs incidentally during transplantation of organs, particularly kidneys or lungs, can lead to subsequent organ deterioration and graft failure.
  • IR ischemia-reperfusion
  • IRI renal ischemia-reperfusion injury
  • the flap refers to the skin or tissue that is moved from one part of the body to another by attaching a vessel and a corresponding tissue that can be viable to the tissue to be moved. Flap is used for soft tissue defects or chronic wounds that cannot be solved by skin grafts, and is the most widely used surgical procedure in the plastic surgery field.It is useful for restoring not only appearance but also loss of function. Due to the complex transplantation of several tissues, such as muscles and nerves, the primary reconstruction is possible, which allows for rapid recovery. In these flaps, the survival rate of the flaps is closely related to the treatment of ischemic reperfusion injury. The ischemia reperfusion injury treatment method, if there is a method that can stably improve the survival rate of the flap is expected to be very useful.
  • ischemia reperfusion injury is an important disease with a high incidence, but there is not enough effective treatment. Therefore, if the effective prevention and treatment for this come out, the ripple effect is likely to be very large.
  • the present inventors have made efforts to develop a composition for treating and preventing ischemic injury, in particular, ischemia reperfusion injury, which has excellent effects, and thus, the present invention has been completed.
  • telomerase a peptide derived from telomerase can have an excellent effect on the treatment and prevention of ischemic damage, in particular ischemic reperfusion injury.
  • composition for treating and preventing ischemic injury comprising a peptide comprising the amino acid sequence of SEQ ID NO: 1, a peptide having a sequence homology of 80% or more with the amino acid sequence, or a fragment thereof.
  • the fragment may be a fragment consisting of three or more amino acids.
  • the ischemic injury is vascular disease, coronary thrombosis, cerebrovascular thrombosis, aneurysm rupture, systemic bleeding, crush injury, sepsis, skin burn, vascular occlusion surgery, cardiopulmonary bypass, organ
  • One or more causes selected from the group consisting of transplantation, cardiopulmonary collapse (acute cardiac death) and asphyxia, and ischemia reperfusion injury.
  • the ischemic injury may be due to ischemia reperfusion injury.
  • the ischemia reperfusion injury is cerebrovascular ischemia reperfusion injury, renal ischemia reperfusion injury, hepatic ischemia reperfusion injury, ischemia reperfusion cardiomyopathy, skin ischemia reperfusion injury, intestinal ischemia reperfusion injury, intestinal ischemia reperfusion injury Injury, gastric ischemia reperfusion injury, pulmonary ischemia reperfusion injury, pancreatic ischemia reperfusion injury, skeletal muscle ischemia reperfusion injury, abdominal muscle ischemia reperfusion injury, limb ischemia reperfusion injury, ischemia reperfusion colitis, mesenteric ischemia reperfusion injury, and asymptomatic ischemia reperfusion injury It may be selected.
  • the ischemia reperfusion injury may be due to organ transplantation.
  • the ischemia reperfusion injury may be due to lung transplantation.
  • the ischemia reperfusion injury may be due to kidney transplant.
  • the ischemia reperfusion injury may occur in the kidney or lung.
  • the ischemia reperfusion injury may occur in the flap.
  • the peptide may be derived from human telomerase.
  • the composition may be a pharmaceutical composition.
  • the composition may be a food composition.
  • ischemic injury characterized in that the above-mentioned composition is administered to a subject in need thereof.
  • compositions comprising the peptides of the present invention can be effectively applied to ischemic damage, in particular ischemic reperfusion injury.
  • urea nitrogen BUN
  • creatine in blood taken 24 hours after ischemia reperfusion.
  • Figure 2 is a photograph showing the result of PAS staining kidney tissue 24 hours after ischemia reperfusion.
  • FIG. 3 is a graph showing the results of renal tissue injury scoring on renal tissue after 24 hours of ischemia reperfusion.
  • Figure 4 is a photograph of the result of evaluating the renal tissue TUNNEL staining 24 hours after ischemia reperfusion.
  • FIG. 5 shows TUNNEL positive cell measurement results of renal tissue evaluated 24 hours after ischemia reperfusion.
  • FIG. 6 shows the results of evaluation of congenital immune cell infiltration by immunohistostaining of F4 / 80 (macrophage maker) and Gr-1 (neutrophil maker) in renal tissue after 24 hours of ischemia reperfusion.
  • FIG. 7 is a graph showing the results of positive cell measurement of F4 / 80 (macrophage maker) and Gr-1 (neutrophil maker) in renal tissue after 24 hours of ischemia reperfusion.
  • 8 to 10 are graphs showing the effect of inhibiting the secretion of inflammatory cytokines in kidney tissues 24 hours after ischemia reperfusion.
  • FIG. 11 is a diagram showing the process of inducing ischemia reperfusion injury to evaluate flap survival.
  • FIG. 12 is a graph showing flap survival measurement results of the PEP1 treated group and the saline treated group 7 days after ischemia reperfusion induction.
  • FIG. 13 is a digital photograph through the ImageJ program.
  • FIG. 14 is a graph showing the result of weighing the lower lobe of the transplanted lung immediately after the experiment after rat lung transplantation, and weighing again after drying in a dryer at 60 ° C. for 24 hours.
  • FIG. 15 is a photograph showing the results of analysis of neutrophil content after instillation and re-extraction of 5 mL of saline into the trachea after rat lung transplantation.
  • the present invention may be variously modified and have various embodiments.
  • the present invention will be described in more detail based on examples. However, these examples are not intended to limit the present invention to the specific embodiments, the present invention is capable of various embodiments and applications based on the description of the claims, and all included within the spirit and scope of the present invention It is to be understood to include the transformations, equivalents, or substitutes. In the following description of the present invention, if it is determined that the detailed description of the related known technology may obscure the gist of the present invention, the detailed description thereof will be omitted.
  • Telomere is a genetic material repeatedly present at the end of a chromosome and is known to prevent damage to the chromosome or binding to another chromosome. Each time a cell divides, the telomeres become slightly shorter. After a certain number of cell divisions, the telomeres become very short, and the cells stop dividing and die. On the other hand, elongation of telomeres is known to prolong cell life. For example, cancer cells secrete an enzyme called telomerase, which prevents telomeres from shortening, so that cancer cells can continue to proliferate without dying. The inventors have found that peptides derived from telomerase are effective in the treatment and prevention of ischemic reperfusion injury and have completed the present invention.
  • a peptide of SEQ ID NO: 1, a peptide that is a fragment of SEQ ID NO: 1, or a peptide having a sequence homology of at least 80% with the peptide sequence is selected from telomerase, specifically human (Homo sapiens) telomerase. Peptides derived.
  • Peptides disclosed herein can include peptides having at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% homology.
  • the peptides disclosed herein, peptides or fragments thereof comprising SEQ ID NO: 1 and one or more amino acids, two or more amino acids, three or more amino acids, four or more amino acids, five or more amino acids, six or more amino acids Or peptides with seven or more amino acids changed.
  • amino acid changes belong to a property that allows the physicochemical properties of the peptide to be altered.
  • amino acid changes can be made, such as improving the thermal stability of the peptide, altering substrate specificity, changing the optimal pH, and the like.
  • amino acid includes not only the 22 standard amino acids that are naturally incorporated into the peptide, but also D-isomers and modified amino acids. Accordingly, in one aspect of the invention the peptide may be a peptide comprising D-amino acids. Meanwhile, in another aspect of the present invention, the peptide may include a non-standard amino acid or the like which has been post-translational modified.
  • post-translational modifications include phosphorylation, glycosylation, acylation (including, for example, acetylation, myristoylation and palmitoylation), alkylation ), Carboxylation, hydroxylation, glycation, biotinylation, ubiquitinylation, changes in chemical properties (e.g., beta-elimination deimidization) , Deamidation) and structural changes (eg, formation of disulfide bridges). It also includes changes in amino acids, such as changes in amino groups, carboxy groups or side chains, caused by chemical reactions that occur during the linkage with crosslinkers to form peptide conjugates.
  • Peptides disclosed herein can be wild-type peptides identified and isolated from a natural source.
  • the peptides disclosed herein may be artificial variants, comprising an amino acid sequence in which one or more amino acids are substituted, deleted and / or inserted compared to peptides that are fragments of SEQ ID NO: 1.
  • Amino acid changes in the wild type polypeptide as well as in artificial variants include conservative amino acid substitutions that do not significantly affect the folding and / or activity of the protein.
  • conservative substitutions include basic amino acids (arginine, lysine and histidine), acidic amino acids (glutamic acid and aspartic acid), polar amino acids (glutamine and asparagine), hydrophobic amino acids (leucine, isoleucine, valine and methionine), aromatic amino acids (phenylalanine, Tryptophan and tyrosine), and small amino acids (glycine, alanine, serine and threonine). Amino acid substitutions that generally do not alter specific activity are known in the art.
  • the most common exchanges are Ala / Ser, Val / Ile, Asp / Glu, Thr / Ser, Ala / Gly, Ala / Thr, Ser / Asn, Ala / Val, Ser / Gly, Tyr / Phe, Ala / Pro, Lys / Arg, Asp / Asn, Leu / Ile, Leu / Val, Ala / Glu, and Asp / Gly, and vice versa.
  • Other examples of conservative substitutions are shown in the following table.
  • Substantial modifications in the biological properties of the peptide include (a) their effect on maintaining the structure of the polypeptide backbone, eg, a sheet or helical conformation, within the substitution region, (b) the charge of the molecule at the target site. Or their effect in maintaining hydrophobicity, or (c) their effect in maintaining the bulk of the side chains, is carried out by selecting significantly different substitutions. Natural residues are divided into the following groups based on common side chain properties:
  • hydrophobic norleucine, met, ala, val, leu, ile
  • Non-conservative substitutions will be made by exchanging a member of one of these classes for another class. Any cysteine residue that is not involved in maintaining the proper conformation of the peptide can generally be substituted with serine to improve the oxidative stability of the molecule and to prevent abnormal crosslinking. Conversely, cysteine bond (s) can be added to the peptide to improve its stability.
  • Another type of amino acid variant of the peptide is a change in the glycosylation pattern of the antibody.
  • change is meant the deletion of one or more carbohydrate residues found in the peptide and / or the addition of one or more glycosylation sites that are not present in the peptide.
  • N-linked refers to a carbohydrate moiety attached to the side chain of an asparagine moiety.
  • Tripeptide sequences asparagine-X-serine and asparagine-X-threonine, where X is any amino acid except proline, are recognition sequences for enzymatic attachment of carbohydrate moieties to asparagine side chains.
  • O-linked glycosylation means attaching one of the sugars N-acetylgalactosamine, galactose or xylose to hydroxyamino acids, most commonly serine or threonine, but 5-hydroxyproline or 5-hydroxylysine You can also use
  • glycosylation sites to the peptide is conveniently performed by changing the amino acid sequence to contain one or more of the above mentioned tripeptide sequences (for N-linked glycosylation sites). Such changes may also be made by adding or replacing one or more serine or threonine residues with the sequence of the original antibody (for O-linked glycosylation sites).
  • ischemic injury refers to damage caused by reduced oxygen transfer while blocking blood circulation in organs that require blood supply such as heart, brain, kidney, and lung, and impaired tissue function and Fatal damage that can result in cell death.
  • Possible causes of ischemic injury include vascular disease, coronary thrombosis, cerebrovascular thrombosis, aneurysm rupture, systemic bleeding, crush injury, sepsis, severe skin burns, vascular occlusion surgery methods (e.g. spinal cord during thoracoabdominal aneurysm surgery) Ischemia), cardiopulmonary bypass method, organ transplantation, cardiopulmonary collapse (acute cardiac death), and asphyxia.
  • ischemic injury encompasses ischemic reperfusion injury that may occur by organ transplantation, etc., in addition to ischemic injury that may normally occur.
  • the ischemia reperfusion injury may include cerebrovascular ischemia reperfusion injury, renal ischemia reperfusion injury, hepatic ischemia reperfusion injury, ischemia reperfusion cardiomyopathy, skin ischemia reperfusion injury, intestinal ischemia reperfusion injury, intestinal ischemia reperfusion injury, gastric ischemia reperfusion injury, lung ischemia reperfusion injury , Pancreatic ischemia reperfusion injury, skeletal muscle ischemia reperfusion injury, abdominal muscle ischemia reperfusion injury, limb ischemia reperfusion injury, ischemia reperfusion colitis, mesenteric ischemia reperfusion injury, and asymptomatic ischemia reperfusion injury, and the like.
  • Ischemic reperfusion injury can occur frequently during organ transplantation.
  • renal transplantation is associated with progressive loss of function and renal impairment of ischemic renal impairment, resulting in activation of the innate immune system. Is known to be one of the important pathogenesis mechanisms.
  • the peptide described in SEQ ID NO: 1 is shown in Table 2 below. "Name” in Table 2 below is named to distinguish peptides.
  • the peptide set forth in SEQ ID NO: 1 represents the entire peptide of human telomerase.
  • a peptide having a sequence of SEQ ID NO: 1, a peptide that is a fragment of SEQ ID NO: 1, or a peptide having at least 80% sequence homology with the peptide sequence corresponds to a peptide included in telomerase.
  • synthetic peptides selected and synthesized at the positional peptides.
  • SEQ ID 2 shows the amino acid sequence of the entire telomerase.
  • ischemic damage comprising a peptide comprising the amino acid sequence of SEQ ID NO: 1 (comprising), a peptide having a sequence homology of 80% or more with the amino acid sequence or a fragment thereof as a active ingredient To provide a composition.
  • the composition for treating and preventing ischemic injury is a peptide comprising an amino acid sequence of SEQ ID NO: 1, a peptide having a sequence homology of 80% or more with the amino acid sequence, or a fragment thereof.
  • composition according to one aspect of the present invention can be applied to all animals including humans, dogs, chickens, pigs, cattle, sheep, guinea pigs or monkeys.
  • the composition is an ischemic reperfusion injury treatment comprising a peptide comprising the amino acid sequence of SEQ ID NO: 1, a peptide having a sequence homology of 80% or more with the amino acid sequence or a fragment thereof as a active ingredient And it provides a preventive pharmaceutical composition.
  • the pharmaceutical composition according to one aspect of the present invention may be administered orally, rectal, transdermal, intravenous, intramuscular, intraperitoneal, intramedullary, intradural or subcutaneous.
  • Formulations for oral administration may be, but are not limited to, tablets, pills, soft or hard capsules, granules, powders, solutions or emulsions.
  • Formulations for parenteral administration may be, but are not limited to, injections, drops, lotions, ointments, gels, creams, suspensions, emulsions, suppositories, patches or sprays.
  • compositions according to one aspect of the invention may include additives such as diluents, excipients, lubricants, binders, disintegrants, buffers, dispersants, surfactants, colorants, flavoring or sweetening agents as needed.
  • additives such as diluents, excipients, lubricants, binders, disintegrants, buffers, dispersants, surfactants, colorants, flavoring or sweetening agents as needed.
  • Pharmaceutical compositions according to one aspect of the invention may be prepared by conventional methods in the art.
  • the active ingredient of the pharmaceutical composition according to one aspect of the present invention will vary depending on the age, sex, weight, pathology and severity of the subject to be administered, the route of administration or the judgment of the prescriber. Dosage determination based on these factors is within the level of ordinary skill in the art, and its daily dose is, for example, 0.1 ⁇ g / kg / day to 1 g / kg / day, specifically 1 ⁇ g / kg / day to 10 mg / kg Per day, more specifically 10 ⁇ g / kg / day to 1 mg / kg / day, and more specifically 50 ⁇ g / kg / day to 100 ⁇ g / kg / day, but is not limited thereto.
  • the pharmaceutical composition according to one aspect of the present invention may be administered once to three times a day, but is not limited thereto.
  • the composition is an ischemic reperfusion injury treatment comprising a peptide comprising the amino acid sequence of SEQ ID NO: 1, a peptide having a sequence homology of 80% or more with the amino acid sequence or a fragment thereof as a active ingredient And prophylactic food compositions.
  • the formulation of the food composition according to one aspect of the present invention is not particularly limited, but may be, for example, formulated into tablets, granules, powders, solutions, solid preparations, and the like.
  • Each formulation may be appropriately selected and formulated by those skilled in the art according to the formulation or purpose of use, in addition to the active ingredient, and may be synergistic when applied simultaneously with other raw materials.
  • the daily dosage of the active ingredient is within the level of those skilled in the art, the daily dosage of which is for example specifically 1 ⁇ g / kg / day to 10 mg / kg / day, more specifically 10 ⁇ g / kg / day To 1 mg / kg / day, and more specifically, 50 ⁇ g / kg / day to 100 ⁇ g / kg / day, but is not limited thereto, and various factors such as age, health condition, and complications of the subject to be administered. It may vary.
  • Preferred embodiments of the invention include the most optimal mode known to the inventors for carrying out the invention. Variations of the preferred embodiments may become apparent to those skilled in the art upon reading the foregoing description. The inventors expect those skilled in the art to make appropriate use of such variations, and the inventors expect the invention to be practiced in a manner different from that described herein. Accordingly, the invention includes all modifications and equivalents of the subject matter referred to in the appended claims, as permitted by patent law. Moreover, any combination of the abovementioned elements within all possible variations is included in the invention unless expressly stated to the contrary or apparently contradictory in context. While the invention has been particularly shown and described with reference to exemplary embodiments, those skilled in the art will understand that various changes in form and detail may be made without departing from the spirit and scope of the invention as defined by the following claims.
  • the peptide (PEP 1) having the sequence shown in SEQ ID NO: 1 is administered to the ischemic reperfusion injury of the kidney, lung, and abdominal muscle skin flap, thereby preventing renal damage and improving flap survival, thereby preventing and treating ischemic injury. To check the effect.
  • PEP 1 The peptide of SEQ ID NO: 1 (hereinafter referred to as "PEP 1") was prepared according to the solid phase peptide synthesis known in the art. Specifically, peptides were synthesized by coupling amino acids from the C-terminus one by one through Fmoc solid phase peptide synthesis (SPPS) using ASP48S (Peptron, Inc., Daejeon, Korea). As follows, the first amino acid at the C-terminus of the peptides was attached to the resin. For example:
  • Coupling reagent is HBTU [2- (1H-Benzotriazole-1-yl) -1,1,3,3-tetamethylaminium hexafluorophosphate] / HOBt [N-Hydroxxybenzotriazole] / NMM [4-Methylmorpholine] It was. Fmoc removal was performed using piperidine in DMF in 20% of DMF.
  • Each peptide was synthesized by repeating a process of reacting the amino acids with each other, washing with a solvent, and then deprotecting the amino acid using the state in which the amino acid protecting group was bound to the solid support.
  • the synthesized peptide was separated from the resin and then purified by HPLC, and confirmed by MS and lyophilized.
  • Ischemia reperfusion induction proceeded as follows.
  • a mouse model with renal ischemia reperfusion injury was obtained by inducing ischemia reperfusion by bilateral clamping for 30 minutes in the kidney pedicle and restoring blood flow by removing the clamp after 30 minutes.
  • the experimental group was divided into three groups: administration group (PEP 1), control group (PBS: no PEP 1 administration), and Sham (no bilateral clamping).
  • PEP 1 was subcutaneously injected at 1000 nmol / kg 30 minutes before and 12 hours after induction of ischemia reperfusion.
  • the experiment was performed by inducing ischemia reperfusion kidney injury using C57BL / 6 mice (8 weeks old) (Charles River Laboratories (Walminton, Mass.).
  • the ischemia reperfusion kidney injury model was used to force the pedicle of the kidney into vascular forceps. ) To stop blood flow, induce ischemia for 28 minutes, and then reperfusion.
  • Peptide PEP 1 was diluted in PBS at a concentration of 1000 nmol / kg and administered intraperitoneally (i.p injection) twice 30 minutes before and 12 hours after ischemia reperfusion.
  • the experimental group was divided into administration group (PEP 1), control group (PBS) and Sham group (group without causing ischemia reperfusion injury and without kidney damage).
  • BUN blood urea nitrogen
  • creatine which are renal toxicity indicators
  • kidney tissues were extracted to produce paraffin blocks to produce immunohistochemistry and Histologic studies (immunohistochemical and histological studies) were conducted, and protein was extracted to measure cytokine (cytokine) levels. Creatine concentration and BUN were measured using an automatic analyzer (Technicon RA-1000; Bayer, Tarrytown, NY).
  • the PEP 1 administration group significantly decreased BUN and creatine values compared to the PBS control group (see FIG. 1).
  • kidney tissues were subjected to peridodic acid-Schiff (PAS) staining (PAS staining with a PAS kit according to the manufacturer's Polysciences, Inc. (Warington, PA, USA) protocol). After staining, renal tissue injury scoring was evaluated by renal tissue injury scoring.
  • the PEP 1 administration group showed a remarkable alleviation of renal tissue damage compared to the PBS control group (see FIGS. 2 and 3).
  • Test Example 4 Inhibitory effect of renal apoptosis
  • TUNEL staining was performed on paraffin kidney sections via a TUNEL staining kit made by Roche Applied Sciences (Indianapolis, IN, USA).
  • Test Example 5 Inhibitory effect of innate immune cell infiltration on renal tissue
  • kidney tissues were assessed for infiltration of congenital immune cells by F4 / 80 (macrophage maker) and Gr-1 (neutrophil maker) immunohistostaining. Specifically, macrophages and neutrophil cell penetration staining were performed using an antibody specific for macrophages (F4 / 80, abcam., Cambridge, Mass.) By an immunochemical method on paraffin-containing sections.
  • the PEP 1-administered group was found to significantly reduce macrophage and neutrophil infiltration into renal tissues (see FIGS. 6 and 7).
  • the PEP 1 administration group significantly reduced IL-6, MCP-1, and levels compared to the PBS control group, but no significant change was observed in TNF- ⁇ (see FIGS. 8 to 10).
  • the protective effect of PEP 1 against ischemic reperfusion kidney damage is renal function (BUN, creatine), renal tissue damage (renal apoptosis), renal tissue infiltration (immune cell infiltration) And renal tissue cytokine (cytokine) secretion inhibition.
  • BUN renal function
  • renal tissue damage renal apoptosis
  • renal tissue infiltration immune cell infiltration
  • renal tissue cytokine cytokine
  • Ischemic reperfusion induction of abdominal muscle skin flap proceeded as follows.
  • Rat models with ischemic reperfusion injury were harvested at 5 cm x 5 cm from the skin of a white rat (Sprague-Dawley Rat, 180 g to 230 g), administered with PEP1 or Saline, and clamped. Ischemia was induced, and after 7 hours, the clamp was removed to induce ischemia reperfusion injury by restoring blood flow (see FIG. 11).
  • the experimental group was divided into three groups: the administration group (PEP 1 treatment group), the control group (saline treatment group: no PEP 1 administration group), and the Sham group (group without causing ischemic reperfusion injury).
  • PEP 1 was injected at a concentration of 10 mg / 500 ⁇ l, and 500 ⁇ l of saline was injected intramuscularly 30 minutes before ischemia reperfusion induction and 1, 2, 3, 4, 5, 7 days after each.
  • Flap Survivability was measured 7 days after induction of ischemia reperfusion. The measurement of flap survival was performed through digital photo analysis through the imageJ program.
  • the lung transplantation model for measuring the preventive effect of PEP1 ischemia reperfusion injury was set as follows.
  • Rats with the same genes were used to fundamentally block the rejection response. Rats were targeted to Sprague-Dawley species with weights of 300 g to 350 g for both donors and recipients. The lungs were then removed from the donor rats and implanted into the recipient rats.
  • PEP1 Lung transplanted rats were divided into four groups as follows. In order to determine the appropriate dose level of PEP1, PEP1 was added at low concentration (5 mg) and high concentration (50 mg), respectively.
  • Test Example 1 Measurement of wet / dry weight ratio
  • the lower lobe of the transplanted lung was cut out and weighed immediately, and then weighed again after drying in a dryer at 60 ° C. for 24 hours.
  • Neutrophil content was analyzed by instillation of 5 mL of normal saline into the trachea and re-extracted.

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Abstract

La présente invention concerne une composition de traitement et de prévention des lésions ischémiques. Plus spécifiquement, la présente invention concerne une composition comprenant un peptide dérivé de la télomérase efficace dans le traitement et la prévention des lésions ischémiques. Un peptide comportant une séquence d'un numéro de séquence ID selon la présente invention, ou un peptide ou un fragment dudit peptide présentant 80 % d'homologie avec ladite séquence, est très efficace dans le traitement et la prévention des lésions ischémiques. Ainsi, la composition comprenant le peptide selon la présente invention peut être appliquée efficacement à des lésions ischémiques, en particulier à des lésions de reperfusion ischémique.
PCT/KR2014/004194 2014-04-30 2014-05-09 Composition de traitement et de prévention des lésions ischémiques Ceased WO2015170790A1 (fr)

Priority Applications (8)

Application Number Priority Date Filing Date Title
PCT/KR2014/004194 WO2015170790A1 (fr) 2014-05-09 2014-05-09 Composition de traitement et de prévention des lésions ischémiques
CN201480080217.6A CN106659149B (zh) 2014-04-30 2014-05-28 用于器官、组织或细胞移植的组合物、试剂盒和移植方法
KR1020167031602A KR102232320B1 (ko) 2014-04-30 2014-05-28 장기, 조직 또는 세포 이식용 조성물, 키트 및 이식 방법
ES14890834T ES2962532T3 (es) 2014-04-30 2014-05-28 Composición para el trasplante de órganos, tejidos o células, kit y procedimiento de trasplante
EP14890834.6A EP3138399B1 (fr) 2014-04-30 2014-05-28 Composition pour une transplantation d'organe, de tissu, ou de cellules, trousse et procédé de transplantation
JP2016565207A JP6466971B2 (ja) 2014-04-30 2014-05-28 臓器、組織又は細胞移植用組成物、キット及び移植方法
PCT/KR2014/004752 WO2015167067A1 (fr) 2014-04-30 2014-05-28 Composition pour une transplantation d'organe, de tissu, ou de cellules, trousse et procédé de transplantation
US15/307,632 US10662223B2 (en) 2014-04-30 2014-05-28 Composition for organ, tissue, or cell transplantation, kit, and transplantation method

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KR20090033878A (ko) * 2006-07-24 2009-04-06 포휴먼텍(주) 허혈성 질환의 완화 및 치료를 위한 약학 조성물 및 그를 전달하기 위한 방법
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